The mechanical stability of the myosin subfragment-2 (S2) was tested with simulated force spectroscopy (SFS) and gravitational force spectroscopy (GFS). Experiments examined unzipping S2, since it required less force than stretching parallel to the coiled coil. Both GFS and SFS demonstrated that the force required to destabilize the light meromyosin (LMM) was greater than the force required to destabilize the coiled coil at each of three different locations along S2. GFS data also conveyed that the mechanical stability of the S2 region is independent from its association with the myosin thick filament using cofilaments of myosin tail and a single intact myosin...

Hypertrophic cardiomyopathy (HCM) is one of the most common genetic cardiac diseases and among the leading causes of sudden cardiac death (SCD) in the young. The cellular mechanisms leading to SCD in HCM are not well known. Prolongation of the action potential (AP) duration (APD) is a common feature predisposing hypertrophied hearts to SCD. Previous studies have explored the roles of inward Na+ and Ca2+ in the development of HCM, but the role of repolarizing K+ currents has not been defined. The objective of this study was to characterize the arrhythmogenic phenotype and cellular electrophysiological properties of mice with HCM, induced by myosin-binding protein C (MyBPC) knockout (KO), and to test the hypothesis that remodeling of repolarizing K+ currents causes APD prolongation in MyBPC KO myocytes...

In myocarditis and dilated cardiomyopathy (DCM) patients the immune system may play an important role in disease progression. In this study, we aimed to identify new antigens as a target for autoimmune response that might play a crucial role in these diseases. Therefore, a peptide-array was used to investigate antibody binding profiles in patients with autoimmune myocarditis or DCM compared to healthy controls and thus to identify disease relevant antigens. To analyze the pathogenicity of the identified antigens, an experimental autoimmune myocarditis (EAM) model was used...

BACKGROUND: Previous data suggest that mitral valve leaflets are elongated in hypertrophic cardiomyopathy (HCM), and mitral valve leaflet elongation may constitute a primary phenotypic expression of HCM. Our objective was to measure the length of mitral valve leaflets by cardiovascular magnetic resonance (CMR) in subjects with HCM caused by a Finnish founder mutation in the myosin-binding protein C gene (MYBPC3-Q1061X), carriers of the same mutation without left ventricular hypertrophy, as well as in unselected consecutive patients with HCM, and respective controls...

Myosin-binding protein C (MyBPC) in the muscle sarcomere interacts with several contractile and structural proteins. Mutations in the cardiac isoform (MyBPC-3) in humans, or animal knockout, are associated with cardiomyopathy. Function of the fast skeletal isoform (MyBPC-2) in living muscles is less understood. This question was addressed using zebrafish models, combining gene expression data with functional analysis of contractility and small-angle x-ray diffraction measurements of filament structure. Fast skeletal MyBPC-2B, the major isoform, was knocked down by >50% using morpholino antisense nucleotides...

β-adrenergic stimulation increases cardiac myosin binding protein C (MyBP-C) and troponin I phosphorylation to accelerate pressure development and relaxation in vivo, although their relative contributions remain unknown. Using a novel mouse model lacking protein kinase A-phosphorylatable troponin I (TnI) and MyBP-C, we examined in vivo haemodynamic function before and after infusion of the β-agonist dobutamine. Mice expressing phospho-ablated MyBP-C displayed cardiac hypertrophy and prevented full acceleration of pressure development and relaxation in response to dobutamine, whereas expression of phosphor-ablated TnI alone had little effect on the acceleration of contractile function in response to dobutamine...

BACKGROUND: Besides its antiarrhythmic action, carvedilol has an activity to suppress cardiac tissue damage. However, it is unknown whether it has any effect on cellular apoptosis and ion channel remodelling. PURPOSE: To know whether carvedilol has any effect on apoptosis and ion channel remodeling of HL-1 cells expressing E334K MyBPC, and comparing it with bisoprolol. METHOD: We examined effects of carvedilol and bisoprolol on the levels of pro- and anti-apoptotic proteins and ion channels as well as apoptosis of HL-1 cells transfected with E334K MyBPC using Western blot and flow cytometry...

Highly organized thick filaments in skeletal muscle cells are formed from ~300 myosin molecules. Each thick-filament-associated myosin molecule is thought to be constantly exchanged. However, the mechanism of myosin replacement remains unclear, as does the source of myosin for substitution. Here, we investigated the dynamics of myosin exchange in the myofibrils of cultured myotubes by fluorescent recovery after photobleaching and found that myofibrillar myosin is actively replaced with an exchange half-life of ~3 h...

Depressive-like behaviors in animals are usually assessed by standardized behavioral tests such as the forced swimming test (FST). However, individual variation in test performance may obscure group differences and thereby hinder the discovery of genes responsible for depression. Few reports have shown the influence of individual variability in identifying the genes associated with depressive-like behaviors. In this study, we conducted microarray analysis to identify genes differentially expressed in the prefrontal cortex (PFC) and cerebellum of rats stratified by FST immobility ratio (% immobility in 5 min) into a control group [immobility ratio: -1 to +1 standard deviation (SD) from the mean] and a depressive group (immobility ratio: +1 to +2 SDs above the mean)...

Besides its antiarrhythmic effect on atrial fibrillation, bepridil protects tissue, yet its effect on apoptosis has never been fully tested. We examine the effect of bepridil on apoptosis of HL-1 cells expressing E334K myosin-binding protein C (MyBPC), a model cell of apoptosis. Bepridil was compared with amiodarone, and its effects on the expression of pro- and anti-apoptotic protein and apoptosis of HL-1 cells expressing mutant E334K MyBPC-green fluorescent protein (GFP) was analyzed using Western blot and a flow cytometer...

Dose-dependent oxidative stress by the anthracycline doxorubicin (Dox) and other chemotherapeutic agents causes irreversible cardiac damage, restricting their clinical effectiveness. We hypothesized that the resultant protein oxidation could be monitored and correlated with physiological functional impairment. We focused on protein carbonylation as an indicator of severe oxidative damage because it is irreversible and results in proteasomal degradation. We identified and investigated a specific high-molecular weight cardiac protein that showed a significant increase in carbonylation under Dox-induced cardiotoxic conditions in a spontaneously hypertensive rat model...

Mendelian cardiomyopathies and arrhythmias are characterized by an important genetic heterogeneity, rendering Sanger sequencing very laborious and expensive. As a proof of concept, we explored multiplex targeted high-throughput sequencing (HTS) as a fast and cost-efficient diagnostic method for individuals suffering from Mendelian cardiac disorders. We designed a DNA capture assay including all exons from 130 genes involved in cardiovascular Mendelian disorders and analysed simultaneously four samples by multiplexing...

BACKGROUND: The formation of contractile myofibrils requires the stepwise onset of expression of muscle specific proteins. It is likely that elucidation of the expression patterns of muscle-specific sarcomeric proteins is important to understand muscle disorders originating from defects in contractile sarcomeric proteins. METHODS: We investigated the expression profile of a panel of sarcomeric components with a focus on proteins associated with a group of congenital disorders...

AIMS: Heart failure (HF) with preserved ejection fraction (HFpEF) is a major cause of morbidity and mortality. Key alterations in HFpEF include increased left ventricular (LV) stiffness and abnormal relaxation. We hypothesized that myofilament protein phosphorylation and function are deranged in experimental HFpEF vs. normal myocardium. Such alterations may involve the giant elastic protein titin, which contributes decisively to LV stiffness. METHODS AND RESULTS: LV tissue samples were procured from normal dogs (CTRL) and old dogs with hypertension-induced LV hypertrophy and diastolic dysfunction (OHT/HFpEF)...

An interplay between growth, glucose regulation and hypertrophic cardiomyopathy (HCM) may exist, but has not been studied in detail. The purpose of this study was to characterize morphometric features, insulin-like growth factor-1 (IGF-1) and glucose metabolism in Maine Coon cats with HCM. Body weight, body condition score (BCS), head length and width, and abdominal circumference were measured in Maine Coon cats >2 years of age. Echocardiography and thoracic radiography (for measurement of humerus length, and fourth and twelfth vertebrae length) were also performed...

The activity of protein phosphatase-1 (PP1) inhibitor-1 (I-1) is antithetically modulated by the cAMP-protein kinase A (PKA) and Ca(2+)-protein kinase C (PKC) signaling axes. β-adrenergic (β-AR) stimulation results in PKA-phosphorylation of I-1 at threonine 35 (Thr35) and depressed PP1 activity, while PKC phosphorylation at serine 67 (Ser67) and/or Thr75 increases PP1 activity. In heart failure, pThr35 is decreased while pSer67 and pThr75 are elevated. However, the role of Ser67/Thr75 phosphorylation in vivo and its effects on Ca(2+)-cycling are not known...

BACKGROUND: Most cases of apparently idiopathic hypertrophic cardiomyopathy (HCM) in children are caused by mutations in cardiac sarcomere protein genes. HCM also commonly occurs as an associated feature in some patients with disorders caused by mutations in genes encoding components of the RAS-mitogen activated protein kinase (MAPK) signaling pathway. Although diagnosis of these disorders is based on typical phenotypic features, the dysmorphic manifestations can be subtle and therefore overlooked...

Cardiac myosin binding protein C (c-MyBPC) is a thick filament protein that is expressed in cardiac sarcomeres and is known to interact with myosin and actin. While both structural and regulatory roles have been proposed for c-MyBPC, its true function is unclear; however, phosphorylation has been shown to be important. In this study, we investigate the effect of c-MyBPC and its phosphorylation on two key steps of the cross-bridge cycle using fast reaction kinetics. We show that unphosphorylated c-MyBPC complexed with myosin in 1:1 and 3:1 myosin:c-MyBPC stoichiometries regulates the binding of myosin to actin (K(D)) cooperatively (Hill coefficient, h) (K(D) = 16...

Myosin binding protein C (MYBPC) is a crucial component of the sarcomere and an important regulator of muscle function. While mutations in different myosin binding protein C (MYBPC) genes are well known causes of various human diseases, such as hypertrophic (HCM) and dilated (DCM) forms of cardiomyopathy as well as skeletal muscular disorders, the underlying molecular mechanisms remain not well understood. A variety of MYBPC3 (cardiac isoform) mutations have been studied in great detail and several corresponding genetically altered mouse models have been generated...